Adsorbent Filter Media For Removal Of Biological Contaminants In Process Liquids

ABSTRACT

Adsorbent filter media particularly suited for removal of biological contaminants in process liquids. A porous fixed bed of adsorbent material is formed, using only a granular adsorbent and a water-insoluble thermoplastic binder. The resulting composite filter allows for a higher amount of adsorbent with smaller adsorbent particles than conventional depth filters. Elimination of cellulose fiber, as well as the elimination of the thermoset binder, results in reduced contamination of the process liquid.

This application is a Continuation of U.S. patent application Ser. No.11/656,184 filed Jan. 22, 2007, which claims priority of U.S.provisional application Ser. No. 60/774,773 filed Feb. 17, 2006, thedisclosures of which are hereby incorporated by reference.

BACKGROUND OF THE INVENTION

Cellulosic depth filters, such as Millistak®+ filters commerciallyavailable from Millipore Corporation, are typically used in theproduction of biopharmaceuticals, as derived from mammalian cell culturefor the purpose of clarifying various crude product fluids. Thesecomposite filters include a layer of tightly structured cellulosic depthmedia, and can be optimized to a specific application, such as retainingcolloidal particles and cell debris or retaining whole cells and largerdebris. They combine sequential grades of media in a single filtercartridge. These filters are most commonly used in polishing orsecondary clarification processes to remove small quantities ofsuspended matter from aqueous product (protein) streams. The primaryfunction of these filters is to protect or extend the service life ofmore expensive downstream separation processes, such as sterilefiltration and affinity chromatography. That is, a common applicationfor these filters is as “prefilters”, protecting downstream equipmentand media from colloidal contaminants and other cell debris. Inaddition, such depth filters are also used for the protection of viralclearance filters by removing trace quantities of agglomerated proteins.

It is also known in the industry that composite depth filters also canretain, to varying degrees, some soluble contaminants commonly found inmammalian cell cultures, such as nucleic acids, host cell proteins,lipids, surfactants, etc. This retention capability for certain solublecontaminants is based on the adsorptive properties of the depth filtermedia.

The filter media typically employed in these depth filters includesrefined cellulose fibers (wood pulp and/or cotton derived), diatomaceousearth, and a water-soluble thermoset resin binder. The diatomaceousearth (a natural form of silica containing trace amounts of varioussilicates) in these composites is typically 40-60% by weight, and isbelieved to be the essential component, adsorbing colloidal sizebiological matter such as cell fragments, organelles and agglomeratedproteins, as well as various soluble biochemicals such as proteins,lipids and nucleic acids.

However, one of the principal drawbacks of the use of these cellulosicdepth filters for the production of parenteral drugs and otherpharmaceuticals is the relatively high level of water-solublecontaminants they release into the system. Indeed, extensivepre-flushing is required to reduce the level of these organic andinorganic contaminants to acceptable levels prior to use. Furthermore,the maximum loading of diatomaceous earth adsorbent within the depthfilter media is limited to about 60% by weight, and the minimum particlesize for the adsorbent to be retained in the fiber matrix is about 10microns.

It is therefore an object of the present invention to reduce oreliminate the release of contaminants from adsorbent filters.

It is another object of the present invention to increase the content orloading of adsorbent in filter media.

It is a further object of the present invention to provide a filter,with smaller adsorbent particles in order to maximize the availablesurface area for adsorption.

Other objects and advantages of the invention will be apparent from thefollowing detailed description and the accompanying drawings.

SUMMARY OF THE INVENTION

The problems of the prior art have been overcome by the presentinvention, which provides adsorbent filter media particularly suited forremoval of biological contaminants in process liquids. A porous fixedbed of adsorbent material is formed, using only a granular adsorbent anda water-insoluble thermoplastic binder. The resulting composite filterallows for a higher amount of adsorbent with smaller adsorbent particlesthan conventional depth filters. Elimination of cellulose fiber, as wellas the elimination of the water-soluble thermoset binder, results inreduced contamination of the process liquid. As a result, extensivepre-flushing is no longer required to reduce extraneous contaminants.Improved media performance is obtained by increasing the content of theadsorbent material, and/or by using smaller adsorbent media to maximizethe available surface area for adsorption.

The resulting composite filter material is substantially or completelydevoid of cellulose and thermoset binder, and can be placed in aclarifying system downstream of a bioreactor and upstream of a sterilefilter. Other applications include pretreatment of cell culture fluidsprior to viral clearance filtration as well as chromatographicseparations.

BRIEF DESCRIPTION OF THE DRAWINGS

FIG. 1 is a graph comparing water permeability of composites of thepresent invention with conventional composites;

FIG. 2 is a graph of pre-filter differential pressure versus throughput;

FIG. 3 is a graph of sterile filter differential pressure versusthroughput; and

FIG. 4 is a chart of conductivity values of effluent passing throughvarious media.

DETAILED DESCRIPTION OF THE INVENTION

The filter media of the present invention includes an adsorbent materialand a water-insoluble thermoplastic binder. This combination can be usedto form a porous, fixed bed of adsorbent material with suitablemechanical properties for the application, including permeability,tensile strength and bending strength. The filter media is particularlyuseful as a depth filter.

Suitable adsorbent materials include diatomaceous earth, silica, porousglass, zeolites, and activated carbon. Diatomaceous earth isparticularly preferred. Additionally, chromatography media, of anyvariety of forms (beads, ground powder, etc.) and surface chemistries(ion exchange, hydrophobic, etc.), can be used as adsorbents in suchporous media. Suitable binders include thermoplastic binders such aspolyolefins, preferably polyethylene, polypropylene or mixtures thereof.The binder is preferably used in bead, powder or fiber form. By properchoice of the binder (in terms of a sufficiently high melting orsoftening point), the media can be autoclaved or otherwise steamsterilized or gamma irradiated to help reduce or eliminate anybiological contaminants therein.

The media fabrication process can depend on the binder form used. Themedia can be prepared by blending the binder with the adsorbentmaterial, followed by fusing the adsorbent particles together such as bypartially melting or softening the binder. For example, polyethylenepowder can be blended dry (such as by shaking/tumbling for severalminutes) with the adsorbent particles, such as diatomaceous earth orsilica beads, in proportions from about 1:1 to about 1:3binder:adsorbent, by weight. The resulting blended material can beplaced in a mold and heated (e.g., in a heated hydraulic press) to asuitable temperature to fuse the adsorbent particles, such as 130° C. to160° C. Diatomaceous earth and silica beads bound into 2-4 mm thick padsusing ultra-high molecular weight polyethylene powder (Mipelonm™),commercially available from Mitsui Chemical, have been formed by thismethod, with an average bead diameter of 20-30 microns. As the materialheats and softens in the press, the compressive force should beperiodically adjusted to maintain a constant force during the heat cycle(about 5 to 10 minutes).

Alternatively, a wet-laid process can be used to form the media,particularly where the binder is in the form of fibers. For example,fine polyethylene fibers (Fybrel™, from Mitsui Chemical) can bedispersed in isopropanol and water, then blended into a slurry withdiatomaceous earth powder in proportions of from about 1:1 to about 1:3binder:adsorbent, by weight. The slurry is transferred to a Buchnerfunnel holding a nominal 1 micron nonwoven support material in its baseto prevent the fibers and adsorbent from passing straight through theperforations of the funnel. The bulk of the liquid is then drawn offthrough a vacuum flask. The formed disk is transferred to an oven fordrying and thermally bonding the adsorbent particles together bypartially melting or softening the polyethylene fibers.

Composite materials constructed as above exhibit very high permeability(high porosity) and low particle retention properties relative to theconventional cellulosic depth filter media conventionally available.Gravity settling of diatomaceous earth particles with polyethylene fiberor powder produces a low-density composite structure with relativelylarge void spaces.

To enhance the separation properties of the diatomaceous earth compositematerials of the present invention, the material should be compacted orcompressed prior to and/or during heating. For example, compressing thefilter media samples for about 30 seconds using a pneumatic press in therange of 100-325 psi was found to be effective. The media samples do notrelax substantially after compression; after heating to fuse thestructure into a monolith, the media maintains its compressed thicknessdimension. The composite can be compressed to about 50-70% of itsoriginal volume depending on composition and compression force.

The application of mechanical compression to the composite materialsalso can be used to regulate water permeability, which can approach thatof standard cellulosic depth filter pads. FIG. 1 illustrates waterpermeability measurements (flow rate relative to pressure differential)of composites made from Celpure™ (Advanced Minerals) diatomaceous earthand Fybrel™ (Mitsui Chemical) polyethylene microfibers, compared toconventional Millistak®+ A1HC depth filter samples. The figure showsthat the water permeabilities of the instant composites are of a rangeconsistent with that of the commercial cellulosic depth filter media.

A third granular or fibrous component may be added to theadsorbent/binder mixture as a means of manipulating the permeability ofthe product to allow for more effective use of the contained adsorbent.The additive may be a functional adsorbent or inert material but must beof a size and quantity to measurably affect the permeability of themedia. Test results have shown that affecting media permeability in thismanner does not substantially compromise (reduce) the adsorbent capacityof the media. Creating selectively larger flow channels within the mediaallows for deeper/broader penetration of the process fluid within theporous matrix that can compensate effectively for the overall loweradsorbent content of the media.

In addition to particle capture, the adsorbent porous monoliths of thepresent invention have significant advantages over conventionalcellulosic adsorbent depth filters, particularly with regard to waterextractibles. Any extractible materials are a serious concern in theproduction of parenteral pharmaceuticals. The conventional cellulosicdepth media is known to have a relatively high extractible loading thatrequires extensive flushing prior to use. The present inventors havedemonstrated that the extractibles that contribute to conductivity(inorganics) do not solely or predominantly derive from the diatomaceousearth. Indeed, the composite materials of the present invention, devoidof cellulose and thermoset binder, result in a reduction in effluentconductivity of 75-90% compared to conventional cellulosic media. Theelimination of cellulose and substitution of the thermoset water-solubleresin binder with a water-insoluble binder such as polyethylene resultin DE-based media having a drastically lower amount of inorganicextractibles. For users of such materials, there is a considerablebenefit in reducing flushing requirements with lower risk of productcontamination.

In general terms, one preferred method for preparing a preferredcomposite filter materials of the present invention is as follows. Ultrahigh molecular weight polyethylene powder, with an average particle sizeof 25 μm, and natural diatomite having a particle size range of 0.2-25μm, are dry mixed, batchwise, in a rotary V-blender that includes aninternal high-speed agitator. The mixture is transferred from theblender to a powder dispenser (or applicator). The applicator dispensesthe mixed powders onto a moving web of porous non-woven polyestermaterial at room temperature at a controlled thickness of up to 0.5inch. Multiple applications of different powder mixtures could beapplied in similar fashion to create a gradient composition adsorbentmedia.

The loose mixed powder layer is then lightly compacted and leveled bycontact with an overhead roller before being heated from below byelectrically-heated plates and simultaneously from above by IR lamps tosoften the polyethylene powder. The temperature of the heated platesescalates along the production path to a final temperature ofapproximately 340° F. The temperature of the powder mixture is held at amaximum of around 340° F. for several minutes before applying anadditional non-woven polyether web on top.

The composite material is then continuously compressed at approximately100 psi to a thickness of 0.10-0.20″ by passing through two heatedcalender rollers also set to a temperature of around 340° F. Thefinished material is then allowed to cool on a metal plate open to theair.

Example 1

Samples of various blends of diatomaceous earth and polyethylenemicrofibers fused into pads of approximately 2-4 mm in thickness weretested in a standard clarification process step related to proteinproduct recovery from mammalian cell cultures. The test was to challengethe adsorbent media with a suspension of E. coli lysate (in buffer)under constant flow conditions while monitoring the pressure rise on themedia sample (rate of plugging) as well as the quality of the effluentrelative to the volume of fluid processed. Effluent or filtrate qualitywas measured by directly filtering the fluid through a 0.2 micronsterilizing grade membrane filter, in this case Durapore® GV. Theseexperimental composite samples were again compared to Millistak®+cellulosic depth filter media.

FIGS. 2 and 3 show the pressure profiles for the adsorbent media samplesand Millistak®+pads and the associated sterile filter profiles.Throughput is reported as the volume of fluid processed relative to thevolume of the depth filter media employed (bed volume).

As shown in the figures, the DE/PE fiber composites were capable ofmatching the tightest or most retentive grade of Millistak®+ DE media(75 grade) in both throughput and retention. The various compositesamples have rates of rise in pressure that lie at or below that of the75DE Millistak®+ media. In addition, the rates or pressure rise on thedownstream sterile filters are all nearly equivalent for the compositesof the invention as compared to 75DE Millistak®+media, indicating acomparable level of particle retention.

Example 2

Samples of diatomaceous earth fused into a fixed-bed pad usingpolyethylene powder (Mipelon™, Mitsui Chemical) were tested for theircapacity to protect a viral retentive membrane, NFP Viresolve 180. Inthis test, the DE/PE composite (approximately 3 mm thick) was challengedwith a solution of polyclonal human IgG protein at a concentration of0.5 gm/L. The Viresolve membrane typically yields a capacity ofapproximately 150 L/m² for this feedstock. Using the Millistak®+ A1HCdepth filter to pre-treat this feedstock for the removal of proteinagglomerates, the Viresolve membrane capacity can be increased to therange of 750-1500 L/m². Two samples of diatomaceous earth (Celpure 25and Celpure 300 (Advanced Minerals)) were blended with the Mipelon PEpowder and formed into 2-3 mm pads after heating (without compression).The DE/PE composite samples were then used to pre-treat the IgGfeedstock and the filtrate was again processed through Viresolve 180 todetermine the effect on membrane capacity.

The Celpure 25 (a fine grade DE) yielded a Viresolve capacity of 440L/m² and the Celpure 300 (a larger coarse grain DE) yielded a Viresolvecapacity of >1000 L/m². These tests indicate that a monolith ofdiatomaceous earth formed with a PE powder binder can provide the samelevel of protection for a viral retentive membrane (by the removal ofprotein agglomerates) as currently provided by cellulosic depth filterssuch as Millistak®+depth filters.

Example 3

To gauge the cleanliness of the composite materials of the invention,material samples were flushed with clean deionized water and theconductivity of the effluent, after a prescribed flush volume, wasmeasured. Conductivity values were taken to represent the level ofsoluble metals present in the filter media. FIG. 4 shows theconductivity values obtained for various DE/PE composite samplesrelative to commercial Millistak®+depth filter samples.

The Millistak®+ DE media is a composite of cellulose and diatomaceousearth plus a water-soluble thermoset resin binder. The CE media containsonly cellulose fiber and binder. It is evident from these measurementsthat those extractibles that contribute to conductivity (inorganics) donot derive predominantly from the diatomaceous earth. Comparing thesevalues to the DE/PE composites tested, there is reduction in effluentconductivity of 75-90%.

Example 4

10-20% of 75-100 micron porous glass beads when added to a 2:1 mixtureof powdered polyethylene (20-30 micron) and diatomite (0.5-10 micron)can reduce the hydraulic permeability of the finished media by 10-30%with no measurable loss in adsorbent capacity as measured by particlecapture and process volume.

1. A composite filter material comprising: a heat fused composite filtermaterial including a diatomaceous earth adsorbent and a water-insolublethermoplastic polyethylene binder heat fused together, and having aratio of the adsorbent to the binder about 1:1 to about 1:3 by weight,wherein the composite filter material is formed by heating the adsorbentmaterial and the binder to a temperature at which the binder at leastpartially melts or softens to thermally bond the adsorbent materialtogether, and the composite filter material is substantially devoid ofcellulose and a thermoset binder.
 2. The composite filter material ofclaim 1, further comprising porous glass beads.
 3. The composite filtermaterial of claim 1, wherein the adsorbent material has a particle sizerange about 0.2 microns to about 25 microns, the binder comprisesultra-high molecular weight polyethylene powder having an averageparticle size about 25 microns, and the porous glass beads have anaverage bead diameter about 20 microns to 30 microns.
 4. The compositefilter material of claim 1, wherein the heat fused composite filtermaterial is compressed to about 50-70% of the original volume of thecomposite filter material prior to and/or during heat fusing togetherthe adsorbent material and the binder.
 5. A porous composite monolithfor removing biological contaminants in process liquids comprising: acompressed heat fused composite porous monolith having an adsorbentchromatography media material, and a water-insoluble thermoplasticbinder dry mixture compressed to about 50-70% of the original volume ofthe composite porous monolith prior to and/or during heat fusingtogether the adsorbent chromatography media material and thethermoplastic binder, the composite filter material being substantiallydevoid of cellulose and a thermoset binder, wherein the thermoplasticbinder includes polyolefins, polyethylene, polypropylene, ultra-highmolecular weight polyethylene, and mixtures thereof, and the ratio ofthermoplastic binder to adsorbent chromatography media material ratioabout 1:1 to about 1:3 by weight.
 6. The composite monolith of claim 5,wherein the adsorbent chromatography media material includes beads,powder, or particles, and the thermoplastic binder includes fibers,microfibers, beads or powder.
 7. The composite monolith of claim 5,further comprising porous glass beads.
 8. The composite monolith ofclaim 5, wherein the thermoplastic binder comprises ultra-high molecularweight polyethylene.
 9. The composite monolith of claim 5, wherein thecomposite is formed by heating the adsorbent chromatography mediamaterial and the binder to a temperature at which the binder at leastpartially melts or softens to thermally bond the adsorbentchromatography media material together.
 10. The composite monolith ofclaim 7, wherein the adsorbent chromatography media material has aparticle size range about 0.2 microns to about 25 microns, the bindercomprises ultra-high molecular weight polyethylene powder having anaverage particle size about 25 microns, and the porous glass beads havean average bead diameter about 20 microns to 30 microns.
 11. Thecomposite monolith of claim 5, having a thickness about 2 mm to about 4mm.
 12. The composite monolith of claim 11, comprises an adsorbentfilter media.
 13. The composite monolith of claim 13, comprises apre-filter.
 14. A biopharmaceutical clarification system, comprising: a)a bioreactor; b) a sterile filter downstream of the bioreactor; and c) acomposite filter media downstream of the bioreactor and upstream of thesterile filter, the composite filter media including a heat fusedcomposite porous monolith having an adsorbent chromatography mediamaterial and a water-insoluble thermoplastic binder dry mixturecompressed to about 50-70% of the original volume of the compositeporous monolith prior to and/or during heat fusing together theadsorbent chromatography media material and the thermoplastic binder,the composite filter media being substantially devoid of cellulose and athermoset binder, wherein the thermoplastic binder includes polyolefins,polyethylene, polypropylene, ultra-high molecular weight polyethylene,and mixtures thereof, and the ratio of thermoplastic binder to adsorbentchromatography media material is about 1:1 to about 1:3 by weight. 15.The biopharmaceutical clarification system of claim 14, furthercomprising an aqueous product stream feedstock from the bioreactor,pretreated through the composite filter media, the resulting filtratebeing processed through the sterile filter.
 16. A method of forming aheat fused composite filter media pad comprising the steps of: a)providing an adsorbent material selected from diatomaceous earth,silica, glass, and a zeolite, and a water-insoluble thermoplastic binderdispersed in water selected from polyolefins, polyethylene,polypropylene, ultra-high molecular weight polyethylene, and mixturesthereof; b) blending the adsorbent material and the binder dispersed inwater in a binder:adsorbent proportions about 1:1 to about 1:3 byweight; c) forming a liquid slurry from the blended adsorbent materialand binder; d) filtering the slurry to draw off the liquid; e) thermallybonding the adsorbent particles together by partially melting orsoftening the binder; and f) forming a heat fused composite filter mediapad substantially devoid of cellulose and a thermoset binder, having athickness about 2 mm to 4 mm.
 17. The process of claim 16, wherein theadsorbent material is diatomaceous earth powder and the binder ispolyethylene fibers.
 18. The process of claim 16, furthering comprisingadding porous glass beads in step (b).
 19. The process of claim 18,wherein the diatomaceous earth has a particle size range about 0.2microns to 25 microns, and the porous glass beads have an average beaddiameter about 20 microns to 30 microns.